Early human development depends on the correct temporal and spatial expression of thousands of genes. The presence of an extra chromosome or a chromosome missing in gametes typically leads to failed fertilization or spontaneous abortions, resulting in phenotypes called 'reduced fertility'. Reduced fertility or infertility may have one of many reasons, but is largely a consequence of advanced age, presence of a structural chromosome abnormality such as a translocation or for familial reasons. Many patients suffering from reduced fertility can now be helped to get the babies they have been hoping for though a combination of in vitro fertilization, which increases the number of embryos available for replacement, and 'Preimplantation Genetic Diagnosis (PGD)', which selects those embryos that are chromosomically normal for replacement. The clinical approach is termed 'Preimplantation Genetic Diagnosis (PGD)' of aneuploidy. PGD is based on interphase cell analysis using fluorescence in situ hybridization (FISH). This approach is sensitive to time constraints and the number of cells available (i.e., 1 or 2). Similarly, for neoplastic analysis or fetal peripheral blood sometimes it is necessary to analyze very small samples of tissue, which will require also interphase analysis of chromosomes. Alternative techniques, such as SKY or CGH, cannot be used effectively for PGD because require either metaphase spreads, which cannot be reliable obtained from one single cell, or need too many- days of hybridization, which is incompatible with regular in vitro fertilization. However, current FISH technology can only detect a limited number of chromosome abnormalities in interphase cells. Thus we propose to combine a novel FISH protocol with Spectral Imaging (SIm) detection to determine the exact number and types of all chromosomes, by using up to 8 uniquely labeled, chromosome-specific DNA probes, in three sequential rounds of hybridization. Our previous experience with FISH suggests that cells can safely go through 3 rounds of FISH without losing much specificity. Thus combining three rounds of hybridizations with 8 different sets of probes per hybridization will allow us analyze all 24 types of chromosomes in a single cell. In a previous SBIR grant (1R43HD3501001A 1) we demonstrated that Sim could work in polar bodies biopsied from oocytes, but that interphase FISH-Sim will solve problems related to chromosome overlap and nuclear condensation that affect chromosome painting in metaphase stage nuclei. In phase-I we propose to develop two of the three sets of probes, and to test that these probes will work with 90% efficiency after three rounds of hybridization in single polar bodies.